Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells.
Saskia KönigManfred FliegaufManuel RhielBodo GrimbacherTatjana I CornuToni CathomenClaudio MussolinoPublished in: Genes (2022)
Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been proposed as a strategy to treat the immunological defect and stabilize the disease, especially in cases with severe lung infections. However, because of the potentially severe side effects associated with allogeneic HSCT, this has been considered only for a few patients. Autologous HSCT represents a safer alternative but it requires the removal of the dominant negative mutation in the patients' cells prior to transplantation. Here, we developed allele-specific CRISPR-Cas9 nucleases to selectively disrupt five of the most common STAT3 dominant negative alleles. When tested ex vivo in patient-derived hematopoietic cells, allele-specific disruption frequencies varied in an allele-dependent fashion and reached up to 62% of alleles harboring the V637M mutation without detectable alterations in the healthy STAT3 allele. However, assessment of the gene expression profiles of the STAT3 downstream target genes revealed that, upon activation of those edited patient cells, mono-allelic STAT3 expression (functional haploinsufficiency) is not able to sufficiently restore STAT3-dependent signaling in edited T cells cultured in vitro. Moreover, the stochastic mutagenesis induced by the repair of the nuclease-induced DNA break could further contribute to dominant negative effects. In summary, our results advocate for precise genome editing strategies rather than allele-specific gene disruption to correct the underlying mutations in AD-HIES.
Keyphrases
- crispr cas
- genome editing
- cell proliferation
- end stage renal disease
- induced apoptosis
- ejection fraction
- newly diagnosed
- genome wide
- cell cycle arrest
- bone marrow
- prognostic factors
- stem cells
- dna methylation
- mesenchymal stem cells
- soft tissue
- gene expression
- low dose
- early onset
- poor prognosis
- hematopoietic stem cell
- case report
- endoplasmic reticulum stress
- cell death
- patient reported outcomes
- signaling pathway
- drug induced
- transcription factor
- patient reported
- genome wide identification
- platelet rich plasma
- pi k akt